Author Contributions:
MG and RR designed the study. RK designed and performed sampling of soil arthropods and sorting of morphospecies. AG identified insect samples. MG did the screening, barcoding, sequencing and analysis of the data. MG and RR wrote the paper.
Figure 1 : A) Rarefaction curve of morphospecies found (in black) showing species richness in the soil arthropod community. Colored lines represent expected number of morphospecies. B) The distribution of three endosymbionts screened across different host arthropod orders.
Figure 2 : Phylogenetic analysis of (A) Wolbachia, (B)Cardinium and (C) Arsenophonus found, shown with some known sequences for better resolution. Wolbachia phylogenetic tree was constructed using MLST data in ClonalFrame with 50% majority rule consensus. Cardinium and Arsenophonus phylogeny was made in MEGA7 using 16S rRNA gene fragment. Dotted nodes represent bootstrap value >50. Wolbachia infections are shown as STs whereas Cardinium and Arsenophonus are labelled with host taxa that they infected. Infections obtained in this study are in BOLD. Brugia malayi, Amoebophilus asiaticus andProteus mirabilis were taken as outgroup for Wolbachia ,Cardinium and Arsenophonus phylogenetic analysis, respectively.
Figure 3 : Association between infected host (left) and endosymbiont (right) phylogeny with (A) Wolbachia A supergroup, (B) Wolbachia B supergroup, (C) Cardinium and (D)Arsenophonus Host phylogeny was constructed by MrBayes using partial CO1 sequences. Black dots represent clade credibility value >70. Phylogenetic relationship between differentWolbachia strains was interpreted through ClonalFrame.Cardinium and Arsenophonus 16S rDNA phylogeny was constructed using MEGA7. Correlation between the two phylogenies suggest phylogenetic incongruence and extensive horizontal transfer across host taxa.
Figure 4 : Maximum likelihood phylogenetic trees of coxA(left), concatenated MLST dataset (centre) and gatB (right) gene made using MEGA7. Black dots represent bootstrap value >50.Wolbachia ST-N2 and ST-560 clustered with B supergroup in concatenated MLST phylogenetic tree whereas in coxA phylogenetic tree, these strains clustered with A supergroup indicating recombination between Wolbachia supergroup A and B. Similarly, ST-552 clustered with F supergroup in concatenated MLST tree, but the individualgatB gene tree shows it to be from A supergroup, indicating recombination between A and F supergroup. These three cases (ST-N2, ST-560, and ST-552) represents between supergroup recombination of gene or gene segment.
Table 1 : Allele profiles of MLST genes for 36 unique host-Wolbachia combinations. (Bold numbers represent new alleles and STs).
Table 2 : Recombination events detected in the WolbachiaMLST sequences. Putative breakpoints indicate concatenated sequences of MLST genes in the order (gatB-coxA-hcpA-ftsZ-fbpA ). p-value was kept at < 0.01.